Role of CD8 in aberrant function of cytotoxic T lymphocytes.

Using H-2Kd-restricted photoprobe-specific cytotoxic T lymphocyte (CTL) clones, which permit assessment of T cell receptor (TCR)-ligand interactions by TCR photoaffinity labeling, we observed that the efficiency of antigen recognition by CTL was critically dependent on the half-life of TCR-ligand complexes. We show here that antigen recognition by CTL is essentially determined by the frequency of serial TCR engagement, except for very rapid dissociations, which resulted in aberrant TCR signaling and antagonism. Thus agonists that were efficiently recognized exhibited rapid TCR-ligand complex dissociation, and hence a high frequency of serial TCR engagement, whereas the opposite was true for weak agonists. Surprisingly, these differences were largely accounted for by the coreceptor CD8. While it was known that CD8 substantially decreases TCR-ligand complex dissociation, we observed in this study that this effect varied considerably among ligand variants, indicating that epitope modifications can alter the CD8 contribution to TCR-ligand binding, and hence the efficiency of antigen recognition by CTL.

Differential reactivity of TCR Vbeta10 alleles to a mouse mammary tumor virus superantigen.

Mouse mammary tumor virus (MMTV) expresses a superantigen (SAg) which plays a critical role in the viral life cycle. We have recently described the new infectious MMTV (SIM) encoding a Vbeta4-specific SAg in mice with a TCR-Vbeta(b) haplotype. We have now compared the SAg activity of this virus in BALB/c mice harboring the TCR-Vbeta(a), TCR-Vbeta(b) or TCR-Vbeta(c) haplotypes which differ by a central deletion in the TCR-Vbeta(a) and TCR-Vbeta(c) locus and by mutations in some of the remaining Vbeta elements. Injection of MMTV (SIM) led to a strong stimulation of Vbeta4+ CD4+ T cells in TCR-Vbeta(b) mice, but only to a weak stimulation of these cells in TCR-Vbeta(a) or TCR-Vbeta(c) mice. A large increase in the percentage of Vbeta10+ cells was observed among CD4+ T cells in mice with the Vbeta(a) or Vbeta(c), but not the Vbeta(b) TCR-Vbeta haplotype. Vbeta10+ cells dominated the response when Vbeta10(a/c) and Vbeta4 subsets were present together. This is the first report of a viral SAg interacting with murine Vbeta10+ cells. Six amino acid differences between Vbeta10(a/c) and Vbeta10(b) could account for the gain of reactivity of Vbeta10(a/c) to the MMTV(SIM) SAg. No mutations were found in the hypervariable region 4 (HV4) of the TCR. Mutations at positions 22 and 28 introduce into Vbeta10(a/c) the same amino acids which are found at these positions in the MMTV(SIM)-reactive Vbeta4. Tridimensional models indicated that these amino acids lie close to HV4 and are likely to be important for the interaction of the SAg with the TCR.

NS4B Self-Interaction through Conserved C-Terminal Elements Is Required for the Establishment of Functional Hepatitis C Virus Replication Complexes.

Hepatitis C virus (HCV) is an important human pathogen, persistently infecting more than 170 million individuals worldwide. Studies of the HCV life cycle have become possible with the development of cell culture systems supporting the replication of viral RNA and the production of infectious virus. However, the exact functions of individual proteins, especially of nonstructural protein 4B (NS4B), remain poorly understood. NS4B triggers the formation of specific, vesicular membrane rearrangements, referred to as membranous webs, which have been reported to represent sites of HCV RNA replication. However, the mechanism of vesicle induction is not known. In this study, a panel of 15 mutants carrying substitutions in the highly conserved NS4B C-terminal domain was generated. Five mutations had only a minor effect on replication, but two of them enhanced assembly and release of infectious virus. Ten mutants were replication defective and used for selection of pseudoreversions. Most of the pseudoreversions also localized to the highly conserved NS4B C-terminal domain and were found to restore replication competence upon insertion into the corresponding primary mutant. Importantly, pseudoreversions restoring replication competence also restored heterotypic NS4B self-interaction, which was disrupted by the primary mutation. Finally, electron microscopy analyses of membrane alterations induced by NS4B mutants revealed striking morphological abnormalities, which were restored to wild-type morphology by the corresponding pseudoreversion. These findings demonstrate the important role of the C-terminal domain in NS4B self-interaction and the formation of functional HCV replication complexes.

Role of hepatocyte wounding on " Plasmodium " liver-stage development and survival

RESUME La première étape primordiale au cycle de vie du Plasmodium dans un hôte mammifère est l'invasion des hepatocytes par des sporozoites. L'infection finale des hepatocytes est précédée de la traversée de plusieurs cellules hôtes, rompant les membranes plasmiques et ayant comme résultat la sécrétion des facteurs cytotoliques dans le micro-environnement. Ce matériel endogène libéré est fortement stimulant/immunogène et peut servir de signal de danger initiant des réponses distinctes dans diverses cellules. De nos jours, le caractère essentiel et salutaire de la migration des sporozoites comme étape d'infection du Plasmodium est vivement controversée. Ainsi, notre étude a visé à caractériser l'effet de l'interaction du parasite avec ses cellules hôtes d'un point de vue immunologique. En particulier, nous avons voulu évaluer l'effet de la perte de matériel cellulaire pendant l'infection de Plasmodium sur les hepatocytes primaires de souris et sur des cultures cellulaires HepG2. Nous avons observé que les facteurs cytotoxiques dérivés des cellules endommagés activent NF-κB - un important régulateur de réponse inflammatoires -dans des cellules voisines des cellules endommagés, qui sont des cellules hôtes potentielles pour l'infection finale du parasite. Cette activation de NF-κB s'est produite peu de temps après l'infection et a mené in vitro et in vivo à une réduction d'infection de façon dépendante du temps, un effet qui a pu être compensé par l'addition de BAY11-7082, un inhibiteur spécifique de NF-κB. De plus, aucune activation de NF-κB avec des parasites SPECT-/-, incapables de traverser les hepatocytes, n'a été observée. Nous avons montré parla suite que l'activation de NF-κB induit l'expression de l'enzyme iNOS dans les hepatocytes, qui est responsable d'une diminution des hepatocytes infectés. En outre, les hepatocytes primaires des souris MyD88-/- n'ont montré ni activation de NF-κB, ni expression d'iNOS lors de l'infection, ce qui suggère la participation des membres de famille du Toll/IL-1 récepteur dans la reconnaissance des facteurs cytosoxiques. En effet, le manque de MyD88 a augmenté significativement l'infection in vitro et in vivo. D'autre part, un rôle bénéfique pour l'activation de NF-κB a été évalué. Les cellules infectées étaient plus résistantes contre l'apoptose induite par Fas (CD95/Apo-1) que les cellules non infectées ou les cellules infectées dans lesquelles NF-κB a été bloqué par BAY11-7082 in vitro. Paradoxalement, l'expression d'iNOS contribue à la protection des cellules infectées contre l'apoptose pax Fas, puisque le traitement avec l'inhibiteur spécifique SMT (S-methylisothiourea) a rendu les cellules infectées plus susceptibles à l'apoptose. Un effet bénéfique additionnel pour le parasite est que la plupart des cellules hôtes traversées présentent des peptides du parasite aux cellules T cytotoxiques spécifiques et peuvent donc réorienter la réaction immune spécifique sur les cellules non infectées. Nous montrons que les cellules hôtes endommagés par la migration du parasite induit l'inflammation, qui limite l'ampleur de l'infection. D'autre part, nos données soutiennent que la survie du parasite Plasmodium dans le foie est assurée par une augmentation de la résistance des hepatocytes contre l'apoptose. SUMMARY The first obligatory step of the Plasmodium life cycle in the mammalian host is the invasion of hepatocytes by sporozoites. Final hepatocyte infection involves the penetration of several host cells, whose plasma membranes are ruptured in the process, resulting in the release of cytosolic factors into the microenvironment. This released endogenous material is highly stimulatory / immunogenic and can serve as a danger signal initiating distinct responses in various cells. To date, it is highly controversial whether sporozoite migration through hepatocytes is an essential and beneficial step for Plasmodium infection. Thus, our study aimed at characterizing the effect of the interaction of the parasite with its host cells from an immunological point of view In particular, we wanted to evaluate the effect of cell material leakage during Plasmodium infection on cultured mouse primary hepatocytes and HepG2 cells. We observed that wounded cell-derived cytosolic factors activate NF-κB - a main regulator of host inflammatory responses - in cells bordering wounded cells, which are potential host cells for final parasite infection. This activation of NF-κB occurred shortly after infection and led to a reduction of infection load in a time dependent manner in vitro and in viva, an effect that could be reverted by addition of the specific NF-κB inhibitor BAY11-7082. In addition, no NF-κB activation was observed when SPECT-/- parasites, which are devoid of hepatocyte traversing properties, were used. We provide further evidence that NF-κB activation causes the induction of inducible nitric oxide synthase (iNOS) expression in hepatocytes, and this is, in turn, responsible for a decrease in Plasmodium-infected hepatocytes. Furthermore, primary hepatocytes from MyD88-/- mice showed no NF-κB activation and iNOS expression upon infection, suggesting a role of the Toll/IL-1 receptor family members in sensing cytosolic factors. Indeed, lack of MyD88 significantly increased infection in vitro and in vivo. In a further complementary series of experiments, we assessed a possible beneficial role for the activation of NF-κB. Infected cells were more resistant to Fas (CD95/Apo-1)-mediated apoptosis than uninfected cells or infected cells in which NF-κB was blocked by BAYl1-7082 in vitro. Paradoxically, iNOS expression contributes to the protection of infected cells from Fas-induced apoptosis, since treatment with the specific iNOS inhibitor SMT (S-Methylisothiourea Sulfate) rendered the infected cells more susceptible to apoptosis. An additional beneficial effect of host cell traversal for the parasite is the fact that mainly traversed cells present parasite-derived peptides to specific cytotoxic T cells and therefore may redirect the specific immune response to uninfected cells. In summary, we have shown that host cells wounded by parasite migration induce inflammation, which limits the extent of parasite infection. In addition, our data support the notion that survival of Plasmodium parasites in the liver is mediated by increasing the resistance of hepatocytes to Fas-induced apoptosis.

Rapid antagonistic coevolution between strains of the social amoeba Dictyostelium discoideum.

Social groups face a fundamental problem of overcoming selfish individuals capable of destroying cooperation. In the social amoeba Dictyostelium discoideum, there is evidence that some clones ('cheaters') contribute disproportionately to the viable spores in a fruiting body while avoiding the dead stalk cell fate. It remains unclear, however, whether this cheating is actually the product of selection. Here, I report the results of an experimental evolution study designed to test whether clones of D. discoideum will evolve resistance to cheating in the laboratory with genetic variation created only through spontaneous mutation. Two strains, one green fluorescent protein (GFP)-labelled and one wild-type, were allowed to grow and develop together before the wild-type strain was removed and replaced with a naïve strain evolving in parallel. Over the course of 10 social generations, the GFP-labelled strain reliably increased its representation in the spores relative to control populations that had never experienced the competitor. This competitive advantage extended to the non-social, vegetative growth portion of the life cycle, but not to pairwise competition with two other strains. These results indicate strong antagonism between strains, mediated by ample mutational variation for cheating and also suggest that arms races between strains in the wild may be common.

Superantigen-induced immune stimulation amplifies mouse mammary tumor virus infection and allows virus transmission.

Endogenous and infectious mouse mammary tumor viruses (MMTVs) encode in their 3' long terminal repeat a protein that exerts superantigen activity; that is, it is able to interact with T cells via the variable domain of the T cell receptor (TCR) beta chain. We show here that transmission of an infectious MMTV is prevented when superantigen-reactive cells are absent through either clonal deletion due to the expression of an endogenous MTV with identical superantigen specificity or exclusion due to expression of a transgenic TCR beta chain that does not interact with the viral superantigen. A strict requirement for superantigen-reactive T cells is also seen for a local immune response following MMTV infection. This immune response locally amplifies the number of MMTV-infected B cells, most likely owing to their clonal expansion. Collectively, our data indicate that a superantigen-induced immune response is critical for the MMTV life cycle.

Hepatitis C virus-mediated mitochondrial dysfunctions are prevented and rescued by cyclophilin inhibition

Alisporivir (Debio-025) is an analogue of cyclosporine A andrepresents the prototype of a new class of non-immunosuppressivecyclophilin inhibitors. In vitro and in vivo studies have shownthat alisporivir inhibits hepatitis C virus (HCV) replication andongoing clinical trials are exploring its therapeutic potential inpatients with chronic hepatitis C. Recent data suggest that theantiviral effect is mediated by inhibition of cyclophilin A whichis an essential host factor in the HCV life cycle. However, alisporiviralso inhibits mitochondrial permeability transition by bindingto cyclophilin D. As HCV is known to affect mitochondrialfunction, we explored the effect of alisporivir on HCV proteinmediatedmitochondrial dysfunction. By the use of inducible celllines, which allow to investigate the effects of HCV polyproteinexpression independent from viral RNA replication and whichrecapitulate the major alterations of mitochondrial bioenergeticsobserved in infectious cell systems, we show that alisporivir preventsHCV protein-mediated cytochrome c redistribution,decrease of cell respiration, collapse of mitochondrial membranepotential, overproduction of reactive oxygen species and mitochondrialcalcium overload. Strikingly, some of the HCV-mediatedmitochondrial dysfunctions could even be rescued byalisporivir. These observations provide new insights into thepathogenesis of HCV-related liver disease and reveal an additionalmechanism of action of alisporivir that is likely beneficialin the treatment of chronic hepatitis C.

Identification of GPx8 as a novel cellular substrate of the hepatitis C virus NS3-4A protease

Background: The hepatitis C virus (HCV) NS3-4A protease is not only an essential component of the viral replication complex and a prime target for antiviral intervention but also a key player in the persistence and pathogenesis of HCV. It cleaves and thereby inactivates two crucial adaptor proteins in viral RNA sensing and innate immunity (MAVS and TRIF) as well as a phosphatase involved in growth factor signaling (TC-PTP). The aim of this study was to identify novel cellular substrates of the NS3-4A protease and to investigate their role in the life cycle and pathogenesis of HCV. Methods: Cell lines inducibly expressing the NS3-4A protease were analyzed in basal as well as interferon- α -stimulated states by stable isotopic labeling using amino acids in cell culture (SILAC) coupled with protein separation and mass spectrometry. Candidates fulfilling strin- gent criteria for potential substrates or products of the NS3-4A protease were further investigated in different experimental sys- tems as well as in liver biopsies from patients with chronic hep- atitis C. Results: SILAC coupled with protein separation and mass spectrometry yielded > 5000 proteins of which 21 can- didates were selected for further analyses. These allowed us to identify GPx8, a membrane-associated peroxidase involved in disulfide bond formation in the endoplasmic reticulum, as a novel cellular substrate of the HCV NS3-4A protease. Cleavage occurs at cysteine in position 11, removing the cytosolic tip of GPx8, and was observed in different experimental systems as well as in liver biopsies from patients with chronic hepatitis C. Further functional studies, involving overexpression and RNA silencing, revealed that GPx8 is a proviral factor involved in viral particle production but not in HCV entry or RNA replica- tion. Conclusions: GPx8 is a proviral host factor cleaved by the HCV NS3-4A protease. Studies investigating the consequences of cleavage for GPx8 function are underway. The identification of novel cellular substrates of the HCV NS3-4A protease should yield new insights into the HCV life cycle and the pathogenesis of hepatitis C and may reveal novel angles for therapeutic inter- vention.

The cyclophilin inhibitor alisporivir prevents hepatitis C virus-mediated mitochondrial dysfunction.

Alisporivir (Debio-025) is an analogue of cyclosporine A and represents the prototype of a new class of non-immunosuppressive cyclophilin inhibitors. In vitro and in vivo studies have shown that alisporivir inhibits hepatitis C virus (HCV) replication, and ongoing clinical trials are exploring its therapeutic potential in patients with chronic hepatitis C. Recent data suggest that the antiviral effect is mediated by inhibition of cyclophilin A, which is an essential host factor in the HCV life cycle. However, alisporivir also inhibits mitochondrial permeability transition by binding to cyclophilin D. Because HCV is known to affect mitochondrial function, we explored the effect of alisporivir on HCV protein-mediated mitochondrial dysfunction. Through the use of inducible cell lines, which allow to investigate the effects of HCV polyprotein expression independent from viral RNA replication and which recapitulate the major alterations of mitochondrial bioenergetics observed in infectious cell systems, we show that alisporivir prevents HCV protein-mediated decrease of cell respiration, collapse of mitochondrial membrane potential, overproduction of reactive oxygen species and mitochondrial calcium overload. Strikingly, some of the HCV-mediated mitochondrial dysfunctions could even be rescued by alisporivir. Conclusion: These observations provide new insights into the pathogenesis of HCV-related liver disease and reveal an additional mechanism of action of alisporivir that is likely beneficial in the treatment of chronic hepatitis C. (HEPATOLOGY 2012).

A V beta 4-specific superantigen encoded by a new exogenous mouse mammary tumor virus.

The superantigen (SAg) expressed by mouse mammary tumor virus (MMTV) has been shown to play an essential role in the course of the viral life cycle. In the present study, we describe a V beta 4-specific SAg encoded by a new exogenous MMTV carried by the SIM mouse strain. This is the first report of a viral or bacterial SAg reacting with mouse V beta 4+ T cells. Injection of MMTV(SIM) into adult BALB/c mice leads to a rapid and strong stimulation of V beta 4+ CD4+ T cells, followed by a slow deletion of these cells. Neonatal exposure to the virus also leads to a progressive deletion of V beta 4+ T cells. In contrast to other strong MMTV SAg, this new SAg requires the presence of major histocompatibility complex class II I-E molecules to be presented efficiently to T cells. Sequence analysis revealed a new predicted amino acid sequence in the C-terminal polymorphic region of this SAg. Furthermore, sequence comparisons to the most closely related SAg with different V beta specificities hint at the specific residues involved in the interaction with the T cell receptor.

Life-history evolution and the polyphenic regulation of somatic maintenance and survival.

Here we discuss life-history evolution from the perspective of adaptive phenotypic plasticity, with a focus on polyphenisms for somatic maintenance and survival. Polyphenisms are adaptive discrete alternative phenotypes that develop in response to changes in the environment. We suggest that dauer larval diapause and its associated adult phenotypes in the nematode (Caenorhabditis elegans), reproductive dormancy in the fruit fly (Drosophila melanogaster) and other insects, and the worker castes of the honey bee (Apis mellifera) are examples of what may be viewed as the polyphenic regulation of somatic maintenance and survival. In these and other cases, the same genotype can--depending upon its environment--express either of two alternative sets of life-history phenotypes that differ markedly with respect to somatic maintenance, survival ability, and thus life span. This plastic modulation of somatic maintenance and survival has traditionally been underappreciated by researchers working on aging and life history. We review the current evidence for such adaptive life-history switches and their molecular regulation and suggest that they are caused by temporally and/or spatially varying, stressful environments that impose diversifying selection, thereby favoring the evolution of plasticity of somatic maintenance and survival under strong regulatory control. By considering somatic maintenance and survivorship from the perspective of adaptive life-history switches, we may gain novel insights into the mechanisms and evolution of aging.

Identification of GPx8 as a novel cellular substrate of the hepatitis C virus NS3-4A protease

Background: The hepatitis C virus (HCV) NS3-4A protease is not only an essential component of the viral replication complex and a prime target for a ntiviral intervention but also a key player i n the persistence and pathogenesis of HCV. It cleaves and thereby inactivates two crucial adaptor proteins in viral RNA sensing and innate immunity (MAVS and TRIF) as well as a phosphatase involved in growth factor signaling (TCPTP). T he aim of this study was to identify novel cellular substrates o f the N S3-4A protease and to investigate their role in the replication and pathogenesis of HCV. Methods: Cell lines inducibly expressing t he NS3-4A protease were analyzed in basal as well as interferon-α-stimulated states by stable isotopic l abeling using amino acids in cell culture (SILAC) coupled with protein separation and mass spectrometry. Candidates fulfilling stringent criteria for potential substrates or products of the NS3-4A protease were further i nvestigated in different experimental systems as well a s in liver biopsies from patients with chronic hepatitis C. Results: SILAC coupled with protein separation and mass spectrometry yielded > 5000 proteins of which 18 candidates were selected for further analyses. These allowed us to identify GPx8, a membrane-associated peroxidase involved in disulfide bond formation in the endoplasmic reticulum, as a n ovel cellular substrate of the H CV NS3-4A protease. Cleavage occurs at cysteine in position 11, removing the cytosolic tip of GPx8, and was observed in different experimental systems as well as in liver biopsies from patients with chronic hepatitis C. Further functional studies, involving overexpression and RNA silencing, revealed that GPx8 is a p roviral factor involved in viral particle production but not in HCV entry or HCV RNA replication. Conclusions: GPx8 is a proviral host factor cleaved by the HCV NS3-4A protease. Studies investigating the consequences of GPx8 cleavage for protein function are underway. The identification of novel cellular substrates o f the HCV N S3-4A protease should yield new insights i nto the HCV life cycle and the pathogenesis of hepatitis C and may reveal novel targets for antiviral intervention.

Anthropogenic disturbance as a driver of microspatial and microhabitat segregation of cytotypes of Centaurea stoebe and cytotype interactions in secondary contact zones.

BACKGROUND AND AIMS: In a mixed-ploidy population, strong frequency-dependent mating will lead to the elimination of the less common cytotype, unless prezygotic barriers enhance assortative mating. However, such barriers favouring cytotype coexistence have only rarely been explored. Here, an assessment is made of the mechanisms involved in formation of mixed-ploidy populations and coexistence of diploid plants and their closely related allotetraploid derivates from the Centaurea stoebe complex (Asteraceae). METHODS: An investigation was made of microspatial and microhabitat distribution, life-history and fitness traits, flowering phenology, genetic relatedness of cytotypes and intercytotype gene flow (cpDNA and microsatellites) in six mixed-ploidy populations in Central Europe. KEY RESULTS: Diploids and tetraploids were genetically differentiated, thus corroborating the secondary origin of contact zones. The cytotypes were spatially segregated at all sites studied, with tetraploids colonizing preferentially drier and open microhabitats created by human-induced disturbances. Conversely, they were rare in more natural microsites and microsites with denser vegetation despite their superior persistence ability (polycarpic life cycle). The seed set of tetraploid plants was strongly influenced by their frequency in mixed-ploidy populations. Triploid hybrids originated from bidirectional hybridizations were extremely rare and almost completely sterile, indicating a strong postzygotic barrier between cytotypes. CONCLUSIONS: The findings suggest that tetraploids are later immigrants into already established diploid populations and that anthropogenic activities creating open niches favouring propagule introductions were the major factor shaping the non-random distribution and habitat segregation of cytotypes at fine spatial scale. Establishment and spread of tetraploids was further facilitated by their superior persistence through the perennial life cycle. The results highlight the importance of non-adaptive spatio-temporal processes in explaining microhabitat and microspatial segregation of cytotypes.

Synergistic action of GA-binding protein and glucocorticoid receptor in transcription from the mouse mammary tumor virus promoter.

B lymphocytes are among the first cells to be infected by mouse mammary tumor virus (MMTV), and they play a crucial role in its life cycle. To study transcriptional regulation of MMTV in B cells, we have analyzed two areas of the long terminal repeat (LTR) next to the glucocorticoid receptor binding site, fp1 (at position -139 to -146 from the cap site) and fp2 (at -157 to -164). Both showed B-cell-specific protection in DNase I in vitro footprinting assays and contain binding sites for Ets transcription factors, a large family of proteins involved in cell proliferation and differentiation and oncogenic transformation. In gel retardation assays, fp1 and fp2 bound the heterodimeric Ets factor GA-binding protein (GABP) present in B-cell nuclear extracts, which was identified by various criteria: formation of dimers and tetramers, sensitivity to pro-oxidant conditions, inhibition of binding by specific antisera, and comigration of complexes with those formed by recombinant GABP. Mutations which prevented complex formation in vitro abolished glucocorticoid-stimulated transcription from an MMTV LTR linked to a reporter gene in transiently transfected B-cell lines, whereas they did not affect the basal level. Exogenously expressed GABP resulted in an increased level of hormone response of the LTR reporter plasmid and produced a synergistic effect with the coexpressed glucocorticoid receptor, indicating cooperation between the two. This is the first example of GABP cooperation with a steroid receptor, providing the opportunity for studying the integration of their intracellular signaling pathways.

A regional Industrial Symbiosis methodology and its implementation in Geneva, Switzerland

The Agenda 21 for the Geneva region is the results from a broad consultation process including all local actors. The article 12 stipulates that « the State facilitates possible synergies between economic activities in order to minimize their environmental impacts » thus opening the way for Industrial Ecology (IE) and Industrial Symbiosis (IS). An Advisory Board for Industrial Ecology and Industrial Symbiosis implementation was established in 2002 involving relevant government agencies. Regulatory and technical conditions for IS are studied in the Swiss context. Results reveal that the Swiss law on waste does not hinder by-product exchanges. Methodology and technical factors including geographic, qualitative, quantitative and economical aspects are detailed. The competition with waste operators in a highly developed recycling system is also tackled.The IS project develops an empirical and systematic method for detecting and implementing by-products synergies between industrial actors disseminated throughout the Geneva region. Database management tool for the treatment of input-output analysis data and GIS tools for detecting potentials industrial partners are constantly improved. Potential symbioses for 17 flows (including energy, water and material flows) are currently studied for implementation.